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Angelica sinensis is a Chinese medicinal herb for treating gynecological and gastrointestinal disorders, and also in conjunction with cancer chemotherapy. Aim of the study In the present study, the cytotoxic and anti-proliferative effects of three main Angelica sinensis phthalides, namely n -butylidenephthalide (BLP), senkyunolide A (SKA) and z -ligustilide (LGT), and their synergy on colon cancer HT-29 cells were investigated. Moreover, the results obtained in both human colon cancer HT-29 and normal colon CCD-18Co cells were compared for the investigation of selectivity. Materials and methods MTT and [ 3 H] thymidine incorporation assays were used for the evaluation of cytotoxic and anti-proliferative effects, respectively. Interactions among phthalides were determined by median-effect analysis. Results All three phthalides dose-dependently decreased cell viability more potently in HT-29 than in CCD-18Co cells. The IC 50 values for inhibition of cell proliferation for SKA, LGT and BLP were 54.17 ± 5.10, 60.63 ± 6.79 and 236.90 ± 18.22 μM, respectively, in HT-29 cells. Angelica sinensis extract demonstrated significant synergy in inhibiting cell proliferation. Conclusions The three phthalides might have anti-cancer potential, yet the phthalides, in combination with other ingredients in Angelica sinensis extract, display significant synergy leading to a stronger anti-tumor effect.

Ethnopharmacological relevance Pyrrolizidine alkaloids (PAs) are a group of heterocyclic phytotoxins present in a wide range of plants. The consumption of PA-containing medicinal herbs or PA-contaminated foodstuffs has long been reported to cause human hepatotoxicity. However, the degrees of hepatotoxicity of different PAs are unknown, which makes it difficult to determine a universal threshold of toxic dose of individual PAs for safe regulation of PA-containing natural products. The aim of the present study is to develop a simple and convenient in vitro model to assess the hepatotoxicity of different PAs. Material and methods Six common cytotoxicity assays were used to evaluate the hepatotoxicity of different PAs in human hepatocellular carcinoma HepG2 cells. Results The combination of MTT and bromodeoxyuridine incorporation (BrdU) assays demonstrated to be a suitable method to evaluate the toxic potencies of various PAs in HepG2 cells, and the results indicated that otonecine-type PA (clivorine: IC 20 =0.013±0.004 mM (MTT), 0.066±0.031 mM (BrdU)) exhibited significantly higher cytotoxic and anti-proliferative effects than retronecine-type PA (retrorsine: IC 20 =0.27±0.07 mM (MTT), 0.19±0.03 mM (BrdU)). While as expected, the known less toxic platyphylline-type PA (platyphylline: IC 20 =0.85±0.11 mM (MTT), 1.01±0.40 mM (BrdU)) exhibited significantly less toxicity. The different cytotoxic and anti-proliferative potencies of various PAs in the same retronecine-type could also be discriminated by using the combined MTT and BrdU assays. In addition, the developed assays were further utilized to test alkaloid extract of Gynura segetum , a senecionine and seneciphylline-containing herb, the overall cytotoxicity of two PAs in the extract was comparable to that of these two PAs tested individually. Conclusion Using the developed in vitro model, the cytotoxicity of different PAs and the extract of a PA-containing herb were investigated in parallel in one system, and their different hepatotoxic potencies were determined and directly compared for the first time. The results suggested that the developed model has a great potential to be applied for the quick screening of the toxicity of PAs and PA-containing natural products.